Circulating tumors cells (CTCs) are shed from the primary tumor or its metastatic sites and recapitulate many of the features of tumor heterogeneity. Importantly, the CTCs number follow closely the progression or regression of the disease. However, CTCs circulate in very low number making it difficult to characterized them using standard blood test procedures.
There is no gold standard to refer to, as there is no optimal platform that has yet permitted detailed molecular and functional characterization of the CTCs. The most widely used methods are positive selection methods based on antigen capture on the surface of CTCs using antibodies bound to magnetic particles. They rely on the universal epithelial marker EpCAM (CD326) present on CTCs but absent from white blood cells. For example, the CellSearch™ (Veridex) method is FDA-approved and uses positive selection by anti-EpCAM antibodies followed by cytokeratin histological confirmation. Non-specific background of leukocytes is eliminated using a CD45 staining. However, this method suffers from several drawbacks, including low sensitivity of detection (less than 40% of patients with chemotherapy naïve advanced cancer are identified) as a minimum of 4 CTCs/mL of blood is necessary to positively identify patients. In addition, due to the destructive nature of the CellSearch™ method, it is not amenable to full isolation and molecular characterization of viable CTCs.
EpCAM antibodies have also been used alternatively to isolate or characterize CTCs. In another example, multiple chambers coated with anti-EpCAM antibodies are used to collect CTCs from peripheral blood. This technique is amenable to enumeration and analysis of around 50 CTCs/mL of blood (total of around 100 to 200 CTCs/chip using 2 to 4 mL of blood). This technology permitted longitudinal monitoring of patients. In a further example, characterization by flow cytometry has also been reported using few mL of collected peripheral blood but its applicability is limited by its sensitivity to detect the low number of cells present in a small blood sample. In still another example, the MagSweeper™ which is an alternate magnetic bead method coated with anti-EpCAM antibodies is another available platform to recuperate CTCs from peripheral blood.
Other approaches, not based on EpCAM expression, have also been used including nucleic acid detection methods and isolation methods based on physical characteristics of CTCs. The former approach based on detection of released DNA or RNA from CTCs lacks specificity and relies on PCR based detection of rare cells among large quantities of white blood cells resulting in many false negative or positive tests. On the other hand, detection and isolation of CTCs based on physical properties has potential value because of its relative simplicity, detection irrespective of cell surface markers, low cost and amenability to high throughput screening. However, such platforms relying on cell size and density are flawed by the inherent heterogeneity of CTCs, which vary widely in size within and between patients. Finally, a number of cell based scanning techniques are also emerging using fiber optic or laser scanning for cytological evaluation. However, these methods will also need further careful validation.
It would be highly desirable to be provided with a method of obtaining circulating tumor cells populations, preferably in a viable form, to allow the further characterization of these populations of cells. It would also be highly desirable to be provided with a method of obtaining other circulating cancer cells populations, preferably in a viable form, to allow the further characterization of these population of cells. These cellular populations could be used in screening assays to identify potential anti-cancer therapeutic agents. Alternatively or in combination, these cellular populations can also be used to provide personalized medicine services to cancer patients.